| BACKGROUNDDepression is a kind of common major mental illness which causes the human mood to be low and the mechanism is not fully clear to all ages and people of all walks of life,which is the result of the interaction of social,psychological and physiological factors,and shows the characteristics of high recurrence rate,high suicide rate and heavy burden.In addition,depression is a disease with a high degree of heterogeneity and symptoms,which can cause multiple systemic and multi-level pathological reactions in the human body,but the current clinical antidepressants are mostly singleacting and targeted drugs,resulting in a number of adverse reactions.However,traditional Chinese medicine has become a hotspot in the research and development of antidepressants because of its mild response and relatively less toxic side effects.It is found that ginkgo biloba extract of traditional Chinese medicine with multi-target and multi-pathway effect has an extremely wide range of drug effects.In our previous studies,Diterpene ginkgolides,one of the main active components of ginkgo biloba extract,was found to have significant antidepressant effect on the metabolic level of animal model,but the pathway and molecular mechanism of its specific action were not clear.At the same time,it is found that it mainly contains ginkgo lactone A,B and K,and there is a lack of research on the depression of ginkgo biloba lactone monomer.Therefore,it is helpful to elucidate the molecular mechanism of antidepressants and to provide support and reference for the research and development of antidepressants by means of molecular biology,the verification of the relevant pathway of the antidepressant effect of ginkgo biloba lactone in transcription and protein expression level.OBJECTIVEFirstly,based on the model of chronic DG intervention,the pathway analysis of hippocampus and prefrontal cortex was carried out,the key pathway was selected for transcription and protein level verification,to further explore the molecular mechanism of the antidepressant effect of DG.Secondly,in order to investigate the effect of ginkgo lactone monomer on the behavior phenotype of C57BL/6J mice,we carried out a series of related behavioral experiments to evaluate the role of ginkgo lactone monomer in mouse related behavior phenotype after the intervention of ginkgo lactone A,B and K in C57BL/6J mice respectively.METHODS1.The drug administration intervention model of Sprague Dawley was constructed,and 4 weeks of aseptic saline or DG were injected into the peritoneal cavity.The effect of DG on the behavior phenotype of SD rats was evaluated by depression-related behavioral experiments.2.The related indexes of hepatic and renal function,blood lipids and blood glucose in the DG group and control group were tested to investigate the drug toxic side effects.3.Through the bioinformatics method to carry on the pathway prediction analysis,selects the key pathway to carry on the RT-qPCR and the western blotting verification.4.The intervention model of ginkgo biloba lactone monomer was constructed in C57BL/6J mice,and 2 weeks of ginkgo lactone A,ginkgo lactone B,ginkgo lactone K were injected continuously in the peritoneal cavity respectively,followed by behavioral experiments in mice 1 weeks and 2 weeks after administration.RESULTS1.There were significant differences in behavioral experiments between the DG group and the control group.The immobility time in the tail suspension test was significantly decreased and the locomotor activity in the center of the open field test was significantly increased in the DG group.2.Compared with the control group,there was no statistical difference in the related indexes of hepatic and renal function in DG group.The results showed that the chronic DG intervention had no obvious drug side effects.3.Ingenuity Pathway Analysis indicates that the hippocampus and prefrontal cortex are associated with the NT3-TrkA and RAS-MAPK pathways.After transcription and protein levels were validated,RT-qPCR results showed a significant increase in the level of expression in the hippocampus and prefrontal lobe of NT3 and TrkA in the NT3-TrkA pathway compared to the control group.In the RAS-MAPK pathway,there was no significant differential gene expression in the hippocampus and prefrontal cortex.In addition,Western blotting results showed that in the NT3-TrkA and RAS-MAPK pathways,the Raf,NT3,Ras and TrkA proteins in the DG group were significantly increased,with only a significant increase in TrkA in the prefrontal cortex.4.In the study of the behavior phenotype of ginkgo lactone monomer in mice,it was found that in the sucrose preference test,ginkgo lactone K could effectively improve the anhedonia.In the open field test,Ginkgo lactone A,B and K can effectively improve the anxiety behavior of mice.And in the elevated plus-maze test,Ginkgo lactone A and B can effectively improve the anxiety behavior of mice,in the degree of improvement,Ginkgo lactone B effect is more significant than Ginkgo lactone A,and ginkgo lactone K has the least effect.In addition,there was no significant change in the immobility time of the tail suspension test and forced swimming test in ginkgo lactone A,B and K,indicating that ginkgo lactone A,B and K had no obvious effect on the desperate behavior of mice.CONCLUSIONThe Chronic DG intervention can effectively improve the desperation and anxiety behavior of rats under the premise that there is no significant effect on hepatic and renal function,blood glucose and blood lipids in rats.At the same time,it may be related to the specificity of brain region to reveal its effect at the molecular biology level,and the effect of DG may be mainly by activating the NT3-TrkA and the RAS-MAPK pathway in hippocampus.In addition,in the preliminary study of ginkgo lactone monomer,it was found that ginkgo lactone K could effectively improve the anhedonia of mice,and ginkgo lactone A,B and K could effectively improve the anxiety behavior of mice.These findings provide a new way of thinking in the study of the antidepressant effect of DG and its monomer,and give some help and support to promote the research and development of antidepressants. |
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